Access to this full-text is provided by Springer Nature.
Content available from Nature Communications
This content is subject to copyright. Terms and conditions apply.
ARTICLE
Space station biomining experiment demonstrates
rare earth element extraction in microgravity
and Mars gravity
Charles S. Cockell 1,11 ✉, Rosa Santomartino 1,11, Kai Finster 2, Annemiek C. Waajen 1, Lorna J. Eades3,
Ralf Moeller4, Petra Rettberg 4, Felix M. Fuchs4,5, Rob Van Houdt 6, Natalie Leys 6, Ilse Coninx6,
Jason Hatton7, Luca Parmitano7, Jutta Krause7, Andrea Koehler7, Nicol Caplin7, Lobke Zuijderduijn7,
Alessandro Mariani8, Stefano S. Pellari8, Fabrizio Carubia8, Giacomo Luciani8, Michele Balsamo8,
Valfredo Zolesi8, Natasha Nicholson1, Claire-Marie Loudon1, Jeannine Doswald-Winkler9, Magdalena Herová9,
Bernd Rattenbacher9, Jennifer Wadsworth10, R. Craig Everroad10 & René Demets7
Microorganisms are employed to mine economically important elements from rocks,
including the rare earth elements (REEs), used in electronic industries and alloy production.
We carried out a mining experiment on the International Space Station to test hypotheses on
the bioleaching of REEs from basaltic rock in microgravity and simulated Mars and Earth
gravities using three microorganisms and a purposely designed biomining reactor. Sphingo-
monas desiccabilis enhanced mean leached concentrations of REEs compared to non-biological
controls in all gravity conditions. No significant difference in final yields was observed
between gravity conditions, showing the efficacy of the process under different gravity
regimens. Bacillus subtilis exhibited a reduction in bioleaching efficacy and Cupriavidus
metallidurans showed no difference compared to non-biological controls, showing the
microbial specificity of the process, as on Earth. These data demonstrate the potential for
space biomining and the principles of a reactor to advance human industry and mining
beyond Earth.
https://doi.org/10.1038/s41467-020-19276-w OPEN
1UK Centre for Astrobiology, School of Physics and Astronomy, University of Edinburgh, Edinburgh, UK. 2Department of Bioscience–Microbiology, Ny
Munkegade 116, Building 1540, 129, 8000 Aarhus C, Denmark. 3School of Chemistry, University of Edinburgh, Edinburgh, UK. 4Radiation Biology
Department, German Aerospace Center (DLR), Institute of Aerospace Medicine, Linder Hoehe, Köln, Germany. 5Institute of Electrical Engineering and
Plasma Technology, Faculty of Electrical Engineering and Information Sciences, Ruhr University Bochum, Bochum, Germany. 6Microbiology Unit, Belgian
Nuclear Research Centre, SCK CEN, Mol, Belgium. 7ESTEC, Keplerlaan 1, 2201 AZ Noordwijk, Netherlands. 8Kayser Italia S.r.l., Via di Popogna, 501, 57128
Livorno, Italy. 9BIOTESC, Hochschule Luzern Technik & Architektur, Lucerne School of Engineering and Architecture, Obermattweg 9, 6052
Hergiswil, Switzerland. 10 Exobiology Branch, NASA Ames Research Center, Moffett Field, CA, USA.
11
These authors contributed equally: Charles S. Cockell,
Rosa Santomartino. ✉email: c.s.cockell@ed.ac.uk
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 1
1234567890():,;
Content courtesy of Springer Nature, terms of use apply. Rights reserved
On Earth, microorganisms play prominent roles in natural
processes such as the weathering of rocks into soils and the
cycling of elements in the biosphere. Microorganisms are
also used in diverse industrial and manufacturing processes1–4,for
example in the process called biomining (or bioleaching)5,6.
Microorganisms can catalyse the extraction of valuable elements
from rocks, such as copper and gold7,8. This process can in some
circumstances reduce the environmentally damaging use of toxic
compounds such as cyanides9,10. These microbial interactions with
minerals are also used to decontaminate polluted soils, in a process
called bioremediation10. Acidophilic iron and sulfur-oxidisers are
often used to biomine economic elements from sulfidic ores, but
heterotrophic microorganisms, including bacteria and fungi, can be
effective in bioleaching in environments with circumneutral or
alkaline pH. These organisms can enable leaching by changing the
local pH in the environment, for example by the release of protons
or organic acids. Alternatively, leaching and sequestration of ele-
ments can occur as a consequence of the release of complexing
compounds11–15.
Of important economic and practical interest are rare earth
elements (REEs), which include the lanthanides, scandium and
yttrium. On account of their physical properties, including
ferromagnetism and luminescence, REEs are used in electronic
devices such as cell phones and computer screens, as well as in
catalysis, metal alloy and magnet production, and many other
high-technology applications. Some REEs are identified as
short-term near-critical elements16, meaning that the demand
will soon outstrip supply. Microorganisms are known to be able
to mobilise REEs. For example, REEs are used as a cofactor in
alcohol dehydrogenases in diverse microbial taxa17,18,andthey
were shown to be essential for the survival of an acidophilic
methanotroph in a volcanic mudpot19.Theabilityofmicro-
organisms to mobilise REEs from rocks has been shown for a
variety of different mineral matrices20,21.
As humans explore and potentially settle in space,
microbe–mineral interactions have been recognised to be impor-
tant, including in biomining22–24. In addition to mining beyond
the Earth, advancing our understanding of microbe–mineral
interactions in space could be applied to: (1) soil formation from
nutrient-poor rocks22, (2) formation of biocrusts to control dust
and surface material in enclosed pressurised spaces25, (3) use of
regolith as feedstock within microbial segments of life support
systems26, (4) use of regolith and microbes in microbial fuel cells
(biofuel)22, (5) biological production of mineral construction
materials27. All of these diverse applications have in common that
they require experimental investigations on how microbes attach
to, and interact with, rock and regolith materials in space envir-
onments. Furthermore, there is a need to know how organisms
alter ion leaching and mineral degradation in altered gravity
regimens, which will occur in any extraterrestrial location.
Altered gravity conditions, such as microgravity, are known
to influence microbial growth and metabolic processes28–30.
Although the capacity of prokaryotes to directly sense gravity
remains a point of discussion, gravity influences sedimentation
and convection in bulk fluids31. By allowing for thermal con-
vection and sedimentation, gravity is thought to affect the
mixing of nutrients and waste, thereby influencing microbial
growth and metabolism32–35. Based on these considerations, we
hypothesised that altered gravity regimens would induce
changes in microbial interactions with minerals, and thus
bioleaching.
In this work, we present the results of the European Space
Agency BioRock experiment, performed on the International
Space Station (ISS) in 2019 to investigate the leaching of
elements from basalt36–38, an analogue for much of the regolith
material on the Moon and Mars, by three species of
heterotrophic microorganisms. The experiment compared bio-
leaching at three different levels of gravity: microgravity,
simulated Mars and terrestrial gravity. Results are reported on
thebioleachingofREEs,demonstratingtheeffectiveuseof
microorganisms in biomining beyond Earth using a minia-
turised space biomining reactor.
Results
REE biomining in space. Data were acquired using the BioRock
biomining reactor, designed for these experiments (Fig. 1)
which contained basaltic rock with known REE composition
(Table 1) and major elements (Supplementary Table 1). REEs
bioleached into solution were measured for all three organisms
(S. desiccabilis,B. subtilis,C. metallidurans) in all three gravity
conditions (microgravity, simulated Mars and Earth gravity)
and for non-biological controls (Fig. 2,SupplementaryFig.1
and Supplementary Table 2). The concentrations of leached
REEs in biological and non-biological condition generally fol-
lowed the trends expected from their abundance in the basaltic
rock (Table 1; Supplementary Table 2). Elements with the
highest abundance (e.g. Ce and Nd) showed the highest leached
concentrations while elements with lowest abundance (Tb, Tm
and Lu) exhibited the lowest concentrations.
Statistical analysis across all three organisms and the three
gravity conditions tested in space showed a significant effect
of the organism (ANOVA: F(2,369) =87.84, p=0.001) on
bioleaching. Post-hoc Tukey tests showed all pairwise compar-
isons between organisms to be significant (p< 0.001). There
was a non-significant effect when gravity conditions were
compared (ANOVA: F(2,369) =0.202, p=0.818). The interac-
tion between gravity and the organism was not significant
(ANOVA: F(4, 369) =1.75, p=0.138).
Statistical analysis was carried out on S. desiccabilis bioleaching.
Comparing the difference between biological samples and the
non-biological controls in each gravity condition for S. desicc-
abilis showed that microgravity was not significant (ANOVA:
F(1,69) =2.43, p=0.124), but significant differences between the
biological experiments and the non-biological controls were
observed in simulated Mars (ANOVA: F(1,83) =14.14, p<
0.0001) and Earth gravity (ANOVA: F(1,83) =24.20, p< 0.0001).
The difference in bioleaching between gravity conditions
was not significant (ANOVA: F(2,123) =1.60, p=0.206) for
S. desiccabilis.
For S. desiccabilis, across all individual REEs and across all
three gravity conditions on the ISS, the organism had leached
111.9% to 429.2% of the non-biological controls (Fig. 3aand
Supplementary Table 3). Student’sttests were used to examine
the concentration of individualREEsbioleachedcomparedto
non-biological controls. Bioleaching was significantly higher
than non-biological controls under simulated Mars and Earth
gravity for individual REEs (p< 0.05, Student’sttest, Supple-
mentary Table 4), except for Pr and Nd which were significantly
higher at the p< 0.1 level, and not significant for Ce in simulated
Mars gravity (p=0.102). For the microgravity condition,
none of individual REE concentrations in the biological
experiment was significantly higher than the non-biological
control (p> 0.05) (Supplementary Table 4). The standard
deviations of the microgravity biological and non-biological
controls for the individual REEs for S. desiccabilis were, apart
from Pr in the biological experiment, higher than for B. subtilis
and C. metallidurans.
Student’sttest comparisons were carried out between the
concentrations of bioleached REEs in different gravities for each
element for S. desiccabilis (Supplementary Table 4). Comparison
between the simulated Mars gravity and simulated Earth gravity
ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w
2NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications
Content courtesy of Springer Nature, terms of use apply. Rights reserved
showed that the concentrations of five elements (La, Sm, Eu, Tb,
Ho) were significantly different at the p< 0.05 level and five more
elements (Gd, Dy, Er, Tm, Yb) at the p< 0.1 level, with simulated
Earth gravity values being higher. These differences were more
evident among the ‘heavy’REEs (elements from Gd up to Lu)
(Fig. 3a). The total quantity of REEs released by S. desiccabilis as a
percentage of the available quantity in the basalt, ranged between
1.17 × 10−1and 2.41 × 10−2% (Supplementary Table 5).
c
d
Filled culture
chambers
a
Body
containing
medium and
fixative
b
Basalt
slide
Gravity vector
42 mm
1.5 cm
Closed
chamber
Medium-filled
chamber
Top
Fig. 1 The BioRock Experimental Unit. a Top-down image of one Experimental Container (EC) containing one EU (Experimental Unit) showing both culture
chambers inflated with medium. bSideways cross section through culture chamber showing location of basalt slide at the back of the chamber and
principle of medium injection and inversion of membrane (shown here in yellow; left side closed, right side inflated with medium). cImage of basalt slide in
a Petri dish submerged in 50% R2A in a ground experiment. dESA astronaut Luca Parmitano inserts an EC into a KUBIK incubator on board the
International Space Station (image credit to ESA).
Table 1 Content of rare earth elements (REEs; reported as μg/g; mean ± standard deviation) in the basalt substrate used in this
experiment and concentrations (total nanograms leached into the chamber fluid volume of 6 mL) at the end of the BioRock
experiment in S. desiccabilis bioleaching chambers and non-biological controls on-board the International Space Station.
S. desiccabilis non-biological control
REE Concentration in basalt (μg/g) Microgravity Mars gravity Earth gravity Microgravity Mars gravity Earth gravity
La 6.81 3.60 ± 1.26 4.96 ± 0.51 3.74 ± 0.51 3.22 ± 2.20 2.56 ± 0.89 1.66 ± 0.23
Ce 13.53 8.85 ± 2.89 9.26 ± 1.94 7.18 ± 0.99 6.45 ± 3.99 5.79 ± 2.06 4.39 ± 1.26
Pr 2.32 1.12 ± 0.43 1.67 ± 0.48 1.07 ± 0.11 0.96 ± 0.64 0.85 ± 0.28 0.48 ± 0.04
Nd 11.57 5.35 ± 2.02 7.89 ± 1.99 5.20 ± 0.47 4.68 ± 3.49 4.28 ± 1.46 2.28 ± 0.24
Sm 3.04 1.44 ± 0.57 2.03 ± 0.36 1.42 ± 0.12 1.13 ± 0.90 1.06 ± 0.37 0.54 ± 0.07
Eu 1.13 0.51 ± 0.16 0.66 ± 0.07 0.53 ± 0.04 0.44 ± 0.25 0.42 ± 0.11 0.27 ± 0.03
Gd 3.67 2.03 ± 0.86 2.93 ± 0.51 2.18 ± 0.13 1.60 ± 1.37 1.36 ± 0.52 0.70 ± 0.10
Tb 0.57 0.42 ± 0.14 0.57 ± 0.08 0.44 ± 0.01 0.30 ± 0.21 0.26 ± 0.07 0.16 ± 0.02
Dy 3.92 2.82 ± 1.00 3.99 ± 0.55 3.08 ± 0.21 1.86 ± 1.43 1.58 ± 0.52 0.92 ± 0.11
Ho 0.80 0.69 ± 0.27 0.98 ± 0.08 0.78 ± 0.08 0.45 ± 0.37 0.36 ± 0.13 0.20 ± 0.03
Er 2.44 2.34 ± 1.01 3.37 ± 0.22 2.75 ± 0.32 1.49 ± 1.26 1.17 ± 0.47 0.64 ± 0.11
Tm 0.29 0.42 ± 0.16 0.58 ± 0.04 0.49 ± 0.06 0.29 ± 0.19 0.24 ± 0.07 0.16 ± 0.01
Yb 2.11 2.44 ± 1.09 3.52 ± 0.36 2.83 ± 0.35 1.47 ± 1.19 1.16 ± 0.44 0.67 ± 0.11
Lu 0.31 0.49 ± 0.20 0.68 ± 0.08 0.57 ± 0.07 0.33 ± 0.22 0.27 ± 0.08 0.18 ± 0.02
(n=3 biologically independent samples with the exception of one non-biological microgravity and non-biological ground control sample which are not included. Full data set in Supplementary Table 2).
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w ARTICLE
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 3
Content courtesy of Springer Nature, terms of use apply. Rights reserved
Identical statistical analysis was carried out for bioleaching
experiments with B. subtilis and C. metallidurans.ForB.
subtilis, the quantity of REEs bioleached was significantly less
than the non-biological controls in microgravity (ANOVA:
F(1,69) =13.05, p< 0.001) and simulated Mars gravity
(ANOVA: F(1,83) =29.55, p< 0.0001), but marginally not
significant in Earth gravity (ANOVA: F(1,83) =3.79, p=
0.055). The difference in the concentrations of REEs bioleached
between gravity conditions was not significant (ANOVA: F
(2,123) =1.45, p=0.240).
ng in container
Ce ng in container
Nd
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
microgravity
Mars gravity
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Earth gravity
Control
Earth gravity
Ground experiment
Control
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
microgravity
Mars gravity
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Earth gravity
Control
Earth gravity
Ground experiment
Control
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
Earth gravity
microgravity
Mars gravity
microgravity
Mars gravity
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Sphingomonas desiccabilis
Bacillus subtilis
Cupriavidus metallidurans
Earth gravity
Control
Earth gravity
Ground experiment
Control
ng in container
La
0123456
0246810 12 14
0246810
ng in container
Tm
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8
ng in container
Lu
00.1
0.2 0.3 0.4 0.5 0.6 0.7 0.8
ng in container
Tb
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8
ISS
ISS
ISS
Fig. 2 Bioleaching and control leaching of the most and least abundant rare earth elements. Concentrations (ng in total chamber liquid) of rare earth
elements (REEs) in each of the experimental flight and ground control samples at the end of the experiment (described in the text) for each of the three
organisms and non-biological controls. The three most (Ce, Nd, La) and least (Tm, Lu, Tb) abundant REEs are shown here (all others in Supplemental
Fig. 1). ISS shows the International Space Station flight experiments. Circles show triplicate measurements (n =3 biologically independent samples. One
non-biological microgravity and non-biological ground control sample were lost and are not shown) and the mean is given as a triangle. Error bars
represent standard deviations.
ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w
4NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications
Content courtesy of Springer Nature, terms of use apply. Rights reserved
For C. metallidurans, the difference between bioleaching and
the non-biological controls was not significant in all three gravity
conditions: microgravity (ANOVA: F(1,69) =2.25, p< 0.138),
simulated Mars (ANOVA: F(1,83) =3.47, p< 0.066), and Earth
gravity (ANOVA: F(1,83) =0.265, p=0.608). The difference in
bioleaching between gravity conditions was not significant
(ANOVA: F(2,123) =0.71, p=0.496).
Comparisons were made for each REE leached into solution in
the biological experiments compared to the non-biological
control for B. subtilis and C. metallidurans and for each separate
gravity condition (t-test). In B. subtilis, for simulated Mars and
Earth gravity, concentrations of bioleached REEs in solution were
significantly lower compared to the non-biological control
(Supplementary Table 4) at the p< 0.05 level, except for Eu,
Gd, Tb, Ho and Lu, which were significantly lower at the p< 0.1
level, and not significant for Ce in the simulated Earth gravity
condition (pvalue =0.378). In C. metallidurans, Tm, Yb and Lu
were statistically lower at the p< 0.1 level in simulated Mars
gravity (Supplementary Table 4).
Comparisons were also made for each REE leached into
solution in the biological experiments between gravity conditions
for B. subtilis and C. metallidurans (t-test). In B. subtilis cultures,
six elements (Dy, Ho, Er, Tm, Yb, Lu) showed a difference at the
p< 0.05 level between microgravity and simulated Mars gravity
and one element (Ce) at the p< 0.05 level between simulated
Mars and Earth gravity. For C. metallidurans cultures, Ce was the
only element that showed a significant difference at the p< 0.01
level between microgravity and simulated Mars gravity. For both
B. subtilis and C. metallidurans, concentrations of elements
leached as a percentage of the total available in the basalt ranged
from 3.22 × 10−2to 4.14 × 10−3% (Supplementary Table 5).
To test whether the REEs were absorbed onto the cell
membrane or within the microbial cell, ICP-MS analyses of
the cell pellets were performed (Supplementary Table 6). The
S. desiccabilis microgravity
S. desiccabilis
B. subtilis
C. metallidurans
Difference with non biological control (%)
Difference with non biological control (%)
S. desiccabilis Mars gravity
S. desiccabilis Earth gravity
B. subtilis microgravity
B. subtilis Mars gravity
B. subtilis Earth gravity
C. metallidurans microgravity
C. metallidurans Mars gravity
C. metallidurans Earth gravity
a
b
La Ce Pr Nd Sm Eu Gd Tb Dy Ho Er Tm Yb Lu
0
100
200
300
400
500
La Ce Pr Nd Sm Gd Tb Ho Er Tm Yb LuEu Dy
0
100
200
300
400
500
600
700
800
Fig. 3 Effects of microorganisms on rare earth element leaching. a Relative (%) difference in mean concentration of leached REEs in the bulk fluid
between biological experiments and non-biological controls showing microgravity, simulated Mars and Earth gravities on the International Space Station for
the three microorganisms. bGround (true Earth gravity control) experiment for the three microorganisms. Standard deviations reported in Supplemental
Table 3, statistics reported in the main text.
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w ARTICLE
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 5
Content courtesy of Springer Nature, terms of use apply. Rights reserved
concentrations of REEs in these samples generally accounted for
less than 5% of the total REEs in the bulk solution in the
biological experiments, with a few exceptions. Notably, Eu was
above 5% in all conditions apart from S. desiccabilis in
microgravity and Mars gravity. ANOVA was used to ascertain
whether the biological enhancement of REEs leached into
solution exhibited by S. desiccabilis was also reflected in the
quantity of REEs bound to cells compared to the two other
organisms. In microgravity, there was a significant difference
between the organisms (ANOVA: F(2,125) =3.98, p=0.021),
but post-hoc Tukey showed that only the S. desiccabilis and B.
subtilis pairwise comparison was significant (p=0.016). There
was no significant difference between organisms in Mars gravity
(ANOVA: F(2,125) =0.466, p=0.629). In Earth gravity, there
was a significant difference (ANOVA: F(2,125) =36.94, p<
0.001) with post-hoc Tukey showing p< 0.001 for all pairwise
comparisons apart from S. desiccabilis and C. metallidurans
(p=0.132). In almost all cases the percentage of total REEs
associated with the S. desiccabilis cell pellets were lower than the
other two organisms (Supplementary Table 6). Thus, there was
no evidence for a systematically higher fraction of REEs in the
S. desiccabilis cell pellets. The concentrations of REEs in the
supernatant produced from washing of the cell pellet was below
the detection limit.
Comparison of the REEs leached into solution between the
different gravity regimens of the non-biological control samples
on the ISS (Figs. 2,3a, Supplementary Fig. 1 and Supplementary
Table 2) showed that the gravity condition was not significant
(ANOVA: F(2,109) =2.91, p=0.059). Student’sttest investiga-
tions of individual elements in each gravity condition (Supple-
mentary Table 4) showed that Pr, Nd, Sm, Eu, Gd, Tb, Dy were
significantly different (at the p< 0.1 level) between simulated
Mars and Earth gravity control samples. The pure 50% R2A
medium and NOTOXhisto fixative contributed low concentra-
tions of REEs (<0.1 ng to the total solution concentration).
S. desiccabilis caused preferential leaching of heavy REEs. The
percentage difference in bioleaching of REEs was calculated for
each microorganism relative to the leaching in the non-biological
controls in the same gravity condition, for space and ground
experiments (Fig. 3and Supplementary Table 3).
S. desiccabilis caused preferential leaching of heavy (Gd up to
Lu) over light (La up to Eu) REEs. On the ISS, the highest
enhancement was a 429.2 ± 92.0% increase in Er leaching in
simulated Earth gravity, compared to the non-biological control.
On the ground, Yb showed the highest enhancement of 767.4 ±
482.4% increase in bioleaching over the non-biological control.
The larger differences between the non-biological and the
biological leaching of heavy REEs compared to light REEs is
reflected in generally lower pvalues (Student’st-tests) for heavy
REEs compared to light REEs (Supplementary Table 4).
Performance of biomining in space and true Earth gravity.In
parallel with the ISS experiment, ground experiments (true Earth
gravity control) were conducted. Results from the ground control
experiments are reported in Figs. 2,3b, Supplementary Fig. 1,
Supplementary Tables 2 and 3. For S. desiccabilis, the effect of the
microorganism on leaching in the ground control compared with
the non-biological control was significant (ANOVA: F(1,68) =
24.56, p< 0.001). All individual elements showed a statistically
significant difference (Student’sttest) with the non-biological
control (Supplementary Table 4) at the p< 0.05 level apart from
two elements at the p< 0.1 level (Nd and Sm) and three elements
with no significant difference (La, Ce, Eu). For B. subtilis, the
effect of the microorganism on leaching was not significant
(ANOVA: F(1,68) =0.034, p=0.854), similarly with C. metal-
lidurans (ANOVA: F(1,68) =0.705, p=0.404).
Bioleaching of REEs in simulated Earth gravity on the ISS
was compared to bioleaching in the ground experiment (true
Earth gravity). S. desiccabilis showed a significant difference
(ANOVA: F(1,82) =8.14, p=0.005) with simulated Earth
gravity on ISS being higher across all REEs. Neither B. subtilis
(ANOVA: F(1,82) =2.42, p=0.124) or C. metallidurans
(ANOVA: F(1,82) =2.45, p=0.121) showed a significant
difference. Non-biological controls exhibited a significant
difference between the simulated Earth gravity on the ISS and
ground controls (ANOVA: F(1,68) =6.90, p=0.011) with the
concentration of REEs leached into solution in simulated Earth
gravity on the ISS being higher across all REEs.
Biomining occurred under near neutral pH conditions. The pH
status is an important factor in the efficacy of biomining.
NOTOXhisto fixative lowers the final pH of the solutions, so that
the pH at the end of the experiment is not representative of the
pH during growth. In all experimental solutions, the final
pH ranged between 4.16 ± 0.20 and 6.12 ± 0.01 (Supplementary
Table 7).
As it was not possible to measure the pH during the
experiment on the ISS, a ground experiment was conducted to
investigate pH changes over the 21 days of growth at 20–22 °C.
Results are shown in Supplementary Fig. 2. The pH remained
circumneutral for the non-biological samples throughout the
experiment with slight differences in the presence of basalt. The
presence of bacteria caused the pH to rise during the 21 days
compared to the negative controls, regardless of the specific
species. At day 21, the pH values for the three cultures in the
presence of basalt were: S. desiccabilis, 8.41 ± 0.01; B. subtilis,
8.63 ± 0.01; C. metallidurans, 8.66 ± 0.01, and the non-biological
control 7.35 ± 0.036 (mean ± sd). The presence of the basalt slide
caused slight pH differences within the biological samples during
the first week of growth. After one week, the pH remained
constant until the end of the experiment for all the microorgan-
isms. After 21 days of growth, the pH values with and without the
presence of the rock are similar for each microorganism,
suggesting that the influence of the rock material on the pH
values stabilised over time (Supplementary Fig. 2). There was a
large drop in pH after the addition of the fixative (S. desiccabilis,
3.58 ± 0.07; B. subtilis, 3.89 ± 0.10; C. metallidurans, 3.76 ± 0.08,
and the non-biological control 3.08 ± 0.03, mean ± sd). The post-
fixative pH values are different depending on the organism, but
independent of the presence of the basalt. After one week of cold
storage, the presence of the basalt slide caused an increase in pH
for all biotic and non-biological samples, indicating that the pH
measured in the flight and ground control samples was influenced
by both the presence of the basalt slide and the fixative.
Discussion
This study investigated the use of microorganisms to extract a
group of economically important elements (fourteen REEs) from
basalt rock, a material found on the Moon and Mars36–38, under
simulated Mars and Earth gravity on the International Space
Station (ISS). Microgravity was investigated as the lowest gravity
level possible to explore the effects of a lack of sedimentation on
bioleaching, to understand the role of gravity in influencing
microbe–mineral interactions in general, and to gain insights into
industrial biomining on asteroids and other very low gravity
planetary objects. A true Earth gravity ground control experiment
was also performed.
The presence of the bacterium S. desiccabilis was found to
enhance mean concentrations of leached REEs in all gravity
ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w
6NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications
Content courtesy of Springer Nature, terms of use apply. Rights reserved
conditions investigated and these enhancements were significant
in simulated Mars and Earth gravity on ISS compared to the non-
biological controls. Although the S. desiccabilis microgravity
samples reached higher mean concentrations than the micro-
gravity non-biological controls for all REEs, the difference was
not statistically significant. The statistical result is interpreted to
be caused by the greater standard deviations in the leached
concentrations of elements in the microgravity biological
experiment and non-biological controls and the loss of one of the
microgravity control samples owing to contamination, rather
than an effect of microgravity on biological leaching.
The lack of a significant difference in the final concentrations
of REEs leached by S. desiccabilis when the different gravity
conditions were compared is surprising since microgravity has
been reported to influence microbial processes39,40. However,
the results are consistent with our observation that final cell
concentrations did not differ between the different gravity
conditions in the three microorganisms31.Onereasonforthe
lack of statistically significant differences in final concentrations
of REEs between gravity conditions might be that the bacterial
cultures had sufficient nutrients to reach their maximum cell
concentration31, regardless of the different sedimentation rates
in each gravity, thus achieving similar leaching concentrations.
Hence, the experiments showed that, with the appropriate
nutrients, biomining is in principle achievable under a wide
range of gravity conditions.
The mechanism for the REE bioleaching in Sphingomonas
desiccabilis is unknown. It was not caused by bulk acidification of
the growth medium, since the ground experiments showed that
the medium had a slightly basic pH profile during the experi-
ment. The microorganism is a prolific producer of extracellular
polysaccharide (EPS) and these compounds are known to
enhance bioleaching in other organisms by complexing ions in
EPS moieties such as uronic acid41,42. A greater biological
enhancement in the leaching of heavy compared to light REEs
was observed, a pattern consistent with observations by Taka-
hashi et al.43 in laboratory cell cultures and natural microbial
biofilms. The authors suggested that phosphate moieties on the
cell or EPS might preferentially bind heavy REEs, a distinct
property of these biologically produced materials. We also note
that the authors suggested that heavy REE enrichments could
potentially be used as a biosignature for the activities of life.
Beyond applications to biomining, our experiments showed the
preferential enhancement of heavy REEs in the liquid phase
including in simulated Martian gravity, indicating the production
of a potential biosignature under altered gravity, with implica-
tions for example for additional methods to test the hypothesis of
life on Mars.
Enhanced REEs associated with pelleted S. desiccabilis cells
compared to the other two species was not observed. The reduced
pH caused during fixation and sample preparation may have
unbound any REEs attached to cell surfaces in all three species.
Alternatively, the majority of the REEs may have bound to the
extracellular EPS or have been released directly into solution. We
have observed S. desiccabilis by confocal microscopy to form
biofilms on the surfaces and at the edges of cavities on the basalt
more pervasively than B. subtilis and C. metallidurans under these
growth conditions, which could have enhanced cell-mineral
interactions and thus leaching of REEs into solution. The analysis
of REEs within biofilms did not form part of this study since we
wished to separately examine the biofilms non-destructively.
Unavoidable in this experiment was the potential for continued
leaching after fixation and during storage, when the pH was
reduced in the chamber. However, during storage, the tempera-
ture was kept at 2.1 °C on the ISS and below 7.1 °C during sample
return to reduce leaching activity44. Furthermore, a similar
reduction of the pH occurred in the non-biological control
samples.
In contrast to S. desiccabilis,B. subtilis demonstrated less mean
leaching in the biological experiments than the non-biological
controls in all three gravity conditions. This cannot be attributed
to cells attached to the rock retarding ion release since the
microorganisms did not form substantial biofilms on the surface
of the rock and the final cell biomass was lower than in the case of
S. desiccabilis31. As the pH was likely to be similar to the other
organisms during the course of the experiment as shown by our
ground-based post-flight pH experiment, differences in pH dur-
ing the experimental phase cannot explain the results. An alter-
native explanation could be a chemical effect of cell exudates,
such as ligands that retarded leaching or the solubility of REEs.
However, despite its previously demonstrated bioleaching
activity45,46, and cell wall absorption of REEs47, Kucuker et al.48
showed that B. subtilis was not able to extract tantalum, a tran-
sition metal considered similar to a REE, from capacitors.
C. metallidurans did not enhance leaching of REEs. In a 3-month
preparatory phase for the BioRock experiments, the leaching of
elements from crushed basalt by this organism on the Russian
FOTON-M4 capsule was investigated49. In this experiment, C.
metallidurans enhanced copper ion release, but other rock elements
did not show significantly enhanced leaching. Although the
microorganism was suspended in mineral water, the results are
consistent with those reported here.
In none of the experiments was a cerium anomaly50 observed.
Unlike other REEs that are all trivalent, cerium can be oxidised to
the less soluble Ce4+state, which can cause differences in pre-
cipitation and concentration compared to other REEs. The
experiments were performed under oxic conditions. However,
once the cerium was leached from the rock, its oxidation state
would not necessarily have changed its presence in the bulk fluid,
potentially explaining the lack of an anomaly.
Comparing the Earth gravity simulation on the ISS with the
ground-based experiments (true 1 × gcontrol), no significant
difference was observed between biological experiments with B.
subtilis and C. metallidurans, but there was a significant differ-
ence between the S. desiccabilis biological experiments and
between the non-biological controls, with ground-based leaching
significantly less in some REEs compared to the Earth gravity
simulation on the ISS. Simulated gravity in space is not exactly
the same as 1 × gon Earth as shear forces induced by cen-
trifugation in space can create different physical conditions.
Furthermore, because of the small radius of the centrifuge rotor
in KUBIK, gravity forces vary across the culture chamber. We
also note that the ground experiment had a 0.46 °C higher tem-
perature offset than the KUBIKs on the ISS during the main
experimental phase. The experiment on the ISS involves the
launch and download to Earth of the samples, which could
influence them in ways that cannot be easily predicted. Never-
theless, the general trends observed in Earth gravity experiments
with respect to biologically enhanced leaching for the three
organisms were conserved in space.
Our experiment has several differences with any proposed
large-scale biomining activity. The basalt rock was not crushed in
order to investigate biofilm formation on a flat, contiguous but
porous rock surface, another main goal of the BioRock experi-
ment. This may have influenced the total percentage of
REEs extracted from the rock, which was generally less than 5 ×
10−2%. These leaching rates would likely be higher with crushed
rocks, which on Earth have been shown to result in leaching
efficiencies of REEs of 8.0 × 10−3% to several tens of percent
under optimised conditions51,52. Furthermore, we did not stir our
reactors as we wanted to investigate the effects of microgravity
and Mars gravity on cell growth in the absence of artificial
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w ARTICLE
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 7
Content courtesy of Springer Nature, terms of use apply. Rights reserved
mixing. Understanding which parameters would require adjust-
ments to enhance the process as well as upscaling of the reactor
would be the next step. Our experiment demonstrates that the
leaching capacities of the three different microorganisms on the
Earth53,54 were similar in space. Thus, Earth-based ground
experiments provide reliable insights into the biomining capa-
cities of specific organisms in space. Yet, our experiments also
confirm that it is important to be careful in the selection of
microorganisms for space biomining operations.
Basaltic material was investigated because it is common on
the Moon and Mars36–38. Our experiment suggests that other
materials could return even higher yields. For example, lunar
KREEP rocks have unusually high concentrations of REEs55,56.
We did not test lunar gravity (0.16 × g) directly, but it lies
between microgravity and Mars gravity. Our results therefore
likely reflect the potential efficacy of biomining operations
under lunar gravity. We suggest the construction of REE bio-
mining facilities in the Oceanus Procellarum and Mare Imbrium
regions of the Moon, where KREEP rocks are abundant. The
principle we demonstrate could be applied to other materials of
economic importance for In-Situ Resource Utilisation (ISRU).
For example, meteoritic material has been shown to be com-
patible with microbial growth26,57–60 and thus our microgravity
experiments show the potential for biomining in low gravity
asteroid environments.
In conclusion, our results demonstrate the biological mining of
economically important elements in space, specifically REEs and
in different extraterrestrial gravity environments. The experi-
ments also demonstrate the novel REE bioleaching ability for the
mesophilic, biofilm-forming, and desiccation-resistant bacterium
S. desiccabilis, which could be used in biomining applications.
From a technical point of view, our experiment also demonstrated
the principles of a miniature space biomining reactor. The
experiment thus shows the efficacy of microbe–mineral interac-
tions for advancing the establishment of a self-sustaining per-
manent human presence beyond the Earth and the technical
means to do that.
Methods
BioRock experiment. BioRock was an experiment proposed to European Space
Agency (ESA) in response to the International Life Science Research Announce-
ment in 2009 (ILSRA-2009). The project was selected as a candidate for flight in
2010 and subsequent bioreactor hardware design has been described61. The
experiment began on the International Space Station on July 30, 2019 and ended on
August 20, 2019.
Microorganisms and growth media. Three bacterial species were used to inves-
tigate bioleaching. Criteria were: (1) they could tolerate desiccation required for
experiment preparation, (2) they could grow on solid surfaces and/or form bio-
films, (3) they were able to interact with rock surfaces and/or bioleach, and (4) they
all could be grown in an identical medium at the same experimental conditions to
allow for comparisons between organisms.
The microorganisms used were:
Sphingomonas desiccabilis CP1D (DSM 16792; Type strain), a Gram-negative,
non-motile, desiccation resistant, non-spore-forming bacterium, which was
isolated from soil crusts in the Colorado plateau62.
Bacillus subtilis NCIB 3610 (DSM 10; Type strain), a Gram-positive, motile,
spore- and biofilm-forming bacterium naturally found in a range of environments,
including rocks63. The organism has been used in several space experiments28,33.
Cupriavidus metallidurans CH34 (DSM 2839; Type strain), a Gram-negative,
motile, non-spore forming bacterium. Strains of this species have been isolated
from metal-contaminated and rock environments64–69. The organism has been
previously used in space experiments70.
The medium used for the BioRock experiment was R2A71 at 50% concentration
as it supported growth of all three microorganisms61, allowing for comparisons.
The composition was (g L−1): yeast extract, 0.25; peptone, 0.25; casamino acids,
0.25; glucose, 0.25; soluble starch, 0.25; Na-pyruvate, 0.15; K
2
HPO
4
, 0.15;
MgSO
4
.7H
2
O, 0.025 at pH 7.2.
NOTOXhisto (Scientific Device Laboratory, IL, USA), a fixative compatible
with safety requirements on the International Space Station (ISS), was used to halt
bacterial metabolism at the end of the experiment.
Bioleaching substrate. Basalt was used for bioleaching, whose REE composition,
as determined by ICP-MS (inductively coupled plasma mass spectrometry) and
bulk composition, as determined by X-ray Fluorescence (XRF), is shown in Table 1
and Supplementary Table 1, respectively. The material was an olivine basalt rock
collected near Gufunes, Reykjavik in Iceland (64°08′22.18′′N, 21°47′21.27′′W)
chosen because it has a chemical composition similar to that of basalts found on
the Moon and Mars36–38. The rock was cut into slides of 1.5 cm × 1.6 cm and 3 mm
thick. The mass of 15 of these slides was 1.871 ± 0.062 g (mean ± standard
deviation). The rock was not crushed, as might be carried out for large-scale
bioleaching, because the BioRock project was also concerned with quantifying the
formation of microbial biofilms over a contiguous mineral surface.
Sample preparation for flight. The basalt rock slides were sterilised by dry-heat
sterilisation in a hot air oven (Carbolite Type 301, UK) for 4 h at 250 °C. This
treatment did not change the mineralogy of the rocks as determined by X-Ray
Diffraction (XRD).
Single strain cultures of each organism were desiccated on the slides as follows:
S. desiccabilis. An overnight culture of the strain was grown in R2A 100% at
20–22 °C until reaching stationary phase (OD600 =0.88 ± 0.09; approximately 109
colony forming units per mL). Then, 1 mL of the culture was inoculated on each
basalt slide and the samples were air-dried at room temperature (≈20–25 °C) with a
sterile procedure within a laminar flow-hood.
B. subtilis. Spores were produced as described previously72. For each basalt slide,
10 µL of a ≈1×10
8spores/mL solution were used as inoculum, i.e. 1 × 106spores
per slide, and air-dried at room temperature (≈20–25 °C) within a laminar flow-
hood.
C. metallidurans. Samples were prepared using a freeze-dry protocol (Belgian
Co-ordinated Collection of Microorganisms, BCCM). Cells were cultured on solid
Tryptone Soya Agar (TSA, Oxoid CM0131, BCCM) medium. When grown
confluently, cells were harvested with a cotton swab and suspended in
cryoprotectant, consisting of sterile horse serum supplemented with 7.5% trehalose
and broth medium no. 2 (625 mg L−1; Oxoid CM0131, BCCM). Thirty millilitres of
bacterial suspension were transferred to a 90 mm petri dish and basalt slides were
submerged in the bacterial suspension and gently shaken overnight. Basalt slides,
each containing approximately 109colony forming units per mL, were then
transferred to a 6-well plate (1 slide per well) and covered with a gas permeable seal
and inserted on a pre-cooled shelf of −50 °C, followed by a freezing phase for
90 min at a shelf temperature of −50 °C. Primary drying was performed with a
shelf temperature of −18 °C and chamber pressure of 400 mTorr. A secondary
drying was performed with a shelf temperature of 20 °C and a chamber pressure
below 10 mTorr. After freeze-dry ing, the 6-well plate was covered with a lid and
wrapped in parafilm.
Negative controls were sterile basalt slides without cell inoculation.
After preparation, all samples were stored at room temperature (20–25 °C) until
integration in the culture chambers in the bioreactor.
Flight experimental setup. The hardware design, assembly and filling procedure
were described previously61. Each Experiment Unit (EU) of the BioRock apparatus
was designed to accommodate two independent basalt slides in two independent
sample chambers (Fig. 1). Each EU contained culture medium and fixative reser-
voirs (Fig. 1a). To allow oxygen diffusion without contaminating the cultures, each
chamber was equipped with a deformable, gas permeable, silicone membrane
(Fig. 1b, c)61. After integration of the basalt slides, the medium and fixative
reservoirs were filled with 5 mL of medium and 1 mL of fixative for each sample,
respectively. The culture chambers and surrounding ducts were purged with
ultrapure sterile N
2
gas.
All the samples were integrated under strict aseptic procedures into the EUs.
There were 36 samples in 18 EUs for the flight experiment, and 12 samples in 6
EUs for the ground experiment. The EUs were integrated into a secondary
container that provided the required two-level containment of the fixative (Fig. 1a).
The EU within the container is referred to as the Experiment Container (EC).
After integration, 18 flight ECs were stored at room temperature (≈20–25 °C)
for 2 days. The ECs were launched to the ISS on board of a Space X Dragon
capsule, Falcon-9 rocket during CRS-18 (Commercial Resupply Services) mission
on July 25, 2019 from the NASA Kennedy Space Center, Cape Canaveral, Florida.
On arrival at the ISS, ECs were stored on-board at 2.1 °C.
On the day of the start of the experiment (July 30, 2019), the ECs were installed
by astronaut Luca Parmitano into two KUBIK facility incubators, pre-conditioned
to a temperature of 20 °C (Fig. 1d). Medium injection was performed robotically,
triggered by internal clocks built within the ECs, powered by electricity provided by
the KUBIK incubator. Thereafter the astronaut removed the ECs and took
photographs of all culture chambers to obtain evidence of the medium supply and
to allow comparison with the same chamber after the experimental growth period.
After image acquisition, the ECs were plugged back into the KUBIKs. Two KUBIK
incubators were used for BioRock, running in parallel: One was set to simulate
Earth gravity (1 × g=9.81 m/s2) at the surface of the basalt slide where bioleaching
is occurring, while the second was set to simulate Mars gravity (0.4 × g=3.71 m/s2;
Mars gravity is strictly 0.38 × g, but finer gresolution is not possible to set in the
KUBIK) at the surface of the basalt slide. Gravity levels were measured every
10 min during the active experimental phase using an accelerometer (ADXL313,
ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w
8NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications
Content courtesy of Springer Nature, terms of use apply. Rights reserved
Analog Devices) mounted on a printed circuit board fixed to the bottom of the EC.
The distance between the top face of the basalt slide and the plane of the top face of
the PCB was 10.3 mm. A correction factor was applied to account for the longer
rotation radius at the basalt slide. These accelerometer (gravity) values are shown
in Supplementary Table 8. The microgravity-exposed ECs were split equally
between both KUBIKs and inserted in the static slots available in the facility. The
experiment was run for 21 days.
To stop the cultures from growing, the fixative was automatically injected into
the culture chambers on August 20, 2019. The samples were removed from the
KUBIK incubators and images of the culture chambers were taken. Afterwards, the
ECs were stored in refrigeration as described below.
Temperature during space experiment. The temperature of the ECs from pre-
flight until post-flight was measured using temperature loggers (Signatrol SL52T
sensors, Signatrol, UK) on the rear of four of the ECs. These data showed that
temperatures did not exceed 7.1 °C from pre-flight handover until storage after
arrival at the ISS. During on-board storage, both before and after the 21-day period
of culturing, temperatures were constant at 2.1 °C. During culturing, the loggers
recorded a temperature of 20.16 °C in both KUBIKs. The ECs were downloaded
from the ISS, packed in a ‘double coldbag’provided by NASA. Splashdown
occurred in the Pacific Ocean on August 27 and handover of the ECs to the
investigators occurred on August 29 at Long Beach Airport, LA, USA. Between
removal from storage on August 26 on ISS and handover to the science team on
August 29, the temperature loggers recorded a temperature of 6.6 °C, rising tran-
siently to 7.1 °C. The ECs were stored in a refrigerated insulated box and trans-
ferred to the NASA Ames Research Centre for sample removal on August 30.
Ground experiment. Parallel to the experiment occurring on the ISS, a 1 × g
ground experiment (true Earth gravity) was run to compare with the ISS simulated
Earth gravity samples. Six ECs for the ground experiment were shipped from the
NASA Kennedy Space Center to the NASA Ames Research Centre under cooled
(4 °C) conditions. The six ECs were attached to a power system (KUBIK Interface
simulation station, KISS) with leads running from the system into a 20 °C
laboratory incubator (Percival E30BHO incubator). The ground reference experi-
ment commenced 2 days after the start of the space experiment and the procedure
for the space experiment was replicated: medium injection, first image acquisition,
21-day experiment, fixation, second image acquisition, and cold storage at 4 °C.
The temperatures of the ECs measured by temperature logger (see above) on two of
the ECs were 3.58 and 4.54 °C during shipment to NASA Ames. During the 21 days
main experimental phase, the loggers recorded a temperature of 20.62 °C. During
post-experiment storage, the temperature was 3.06 °C.
Sample recovery. Liquid and basalt slide removal from the ECs was performed at
the NASA Ames Research Center. From the total of 6 mL of total bulk fluid per EC,
an aliquot of 3 mL was taken and 65% nitric acid was added to a final concentration
4% to fix ions and minimise attachment and loss to container walls. These samples
were cold stored at 4 °C until further analysis.
Fixative injection was successful for all the space ECs. However, fixative
injection failed in four of the ground experiment chambers: one B. subtilis
chamber, two C. metallidurans chamber and one non-biological control sample. In
these cases, 1 mL of NOTOXhisto was added to the liquid samples before the
abovementioned procedures.
In all ECs, two culture chambers were observed to have contamination: an ISS
non-biological control chamber in microgravity, juxtaposed to a B. subtilis
chamber, was contaminated with cells that were morphologically identical to B.
subtilis. In the ground control samples, a non-biological control chamber,
juxtaposed to a B. subtilis chamber, had a cellular contaminant at low
concentration that formed a white pellet on centrifugation that was
morphologically dissimilar to B. subtilis. NOTOXhisto fixation prevented
successful DNA extraction and identification in both cases. These data points were
removed from the calculations.
All samples were shipped back to the University of Edinburgh in cold storage by
Altech Space (Torino, Italy).
ICP-MS analysis of samples. Upon return to Edinburgh, UK, the 3 mL of acid-
fixed sample was prepared in the following way: each sample was sequentially
(in three batches) spun down in a 1.5 mL tube at 10,000 × g(IEC MicroCL 17
centrifuge, Thermo Scientific) for ten minutes to pellet cells and cell debris. The
supernatant was collected into a 15 mL tube and analysed by ICP-MS (inductively
coupled plasma mass spectrometry). Acquired liquids were used to determine the
bulk fluid REE concentrations. Cell debris pellets were washed two times in ddH
2
O
and this discarded liquid was pooled. Nitric acid was added to the pooled fluid to a
final concentration of 4%, and the samples were analysed by ICP-MS. This liquid
was used to determine the REE concentrations that were washed off the cell matter.
The pellet was transferred to an acid-washed glass serum bottle pre-baked at 450 °C
in an oven (Carbolite Type 301, UK) for 4 h to remove organic molecules. The vial
with the pellet was heated at 450 °C for a further 4 h to volatilise carbon and leave
residual ions. After cooling, 1.5 mL of ddH
2
O were added with nitric acid to a final
concentration of 4% and samples were analysed by ICP-MS. This liquid was used
to determine the REE concentrations associated with the cell material.
ICP-MS analysis was carried out as described below on the R2A 50%,
NOTOXhisto and ddH
2
O. It was not possible to examine the separated
cryoprotectant for C. metallidurans. However, as a significance enhancement in the
biological experiments compared to the controls for this organism was not
observed, we infer that the protectant did not add additional REEs.
All samples were analysed by ICP-MS using an Agilent 7500ce (with octopole
reaction system), employing an RF (radio-frequency) forward power of 1540 W
and reflected power of 1 W, with argon gas flows of 0.81 L/min and 0.20 L/min for
carrier and makeup flows, respectively. Sample solutions were taken up into the
micro mist nebuliser by peristaltic pump at a rate of approximately 1.2 mL/min.
Skimmer and sample cones were made of nickel.
The instrument was operated in spectrum multi-tune acquisition mode and
three replicate runs per sample were employed. The isotopes: 139La, 140Ce, 141Pr,
146Nd, 147Sm. 153Eu, 157Gd, 159Tb. 163Dy, 165Ho, 166Er, 169Tm, 172Yb, 175Lu were
analysed in ‘no gas’mode with each mass analysed in fully quantification mode and
three points per unit mass. The REE Pm (promethium) was not measured as the
element is radioactive and no standard was available.
To calibrate the instrument, multi-element calibration standards containing
each element were prepared using an REE multi-element standard (Multi-Element
Calibration Standard-1, Agilent Technologies, USA) plus a Uranium single-
element 1000 mg L−1standard (SPE Science, Canada) diluted with 2% v/v HNO
3
(Aristar grade, VWR International, United Kingdom). A NIST standard reference
material, SRM1640a, was employed as a reference standard for some of the
elements. The limits of detection for the REEs were split broadly into two groups:
La, Ce, Pr, Tb, Ho, Tm, Lu: 0.0025-0.005 ppb. Nd, Sm, Eu, Gd, Dy, Er, Yb: 0.001-
0.005 ppb.
Raw ICP-MS data (determined in μg/L) was converted to obtain the absolute
quantity of a given element in the culture chamber, taking into account dilution
factors applied during ICP-MS analysis.
To determine REE concentrations in the basalt slide, between 25 and 50 mg of
homogenised sample was added to Savillex Teflon vessels. Rock standards (basalt
standards BIR-1, BE-N, BCR-2, BHVO-1) were prepared in the same way. Two
blanks were included (i.e., sample without basalt). Three millilitres of double
distilled HNO
3
, 2 mL HCl and 0.5 mL HF was added to each of the vessels. HF was
added after the other acids to prevent disassociation, formation and precipitation of
aluminium fluorides. Samples were placed on a hot plate for digestion overnight
(temperature of 100–120 °C) and checked for complete digestion. Samples were
evaporated on the hot plate. Five millilitres of 1 M HNO
3
was added to each vessel.
Lids were added and the samples returned to the hot plate for a second digestion
step. Samples were further diluted with 2–5% HNO
3
for ICP-MS analysis.
Analysis was carried out on a high resolution, sector field, ICP-MS (Nu AttoM).
The ICP-MS measurements for REEs were performed in low resolution (300), in
Deflector jump mode with a dwell time of 1 ms and 3 cycles of 500 sweeps. Data
were reported in micrograms of REE per gram of basalt.
pH of flight experiment and ground pH experiment. A small aliquot (≈0.3 mL)
of liquid from the chambers was used to measure the pH of the solutions at the end
of the experiment after fixative addition. The pH was measured using a calibrated
Mettler Toledo Semi-Micro-L pH metre. Final values for cell growth in the
experiment are reported previously31 and the values are shown in Supplementary
Table 9.
During the space experiment, only final pH values were obtained. Thus, an
experiment was carried out on the ground to investigate the pH changes that might
have occurred during the course of the experiment (limited to a 1 × gcondition)
and the influence of the basalt rock in any observed pH changes.
Sterile basalt slides, as used in the flight experiment, were prepared in 5 mL of
50% R2A in six-well plates (Corning, UK) and the wells were inoculated with one
of the three microorganisms used in this study. Control experiments were
conducted without organisms, using fresh 50% R2A only, with or without the
basalt slide. The experiment was performed at 20 °C for 21 days. After this period,
1 mL of NOTOXhisto was added to each well, and stored at 4 °C for a further week.
During the course of the experiment, pH values were measured at fixed intervals
(day 0, 1, 4, 7, 14, 21). On the 21st day of the experiment, pH was measured twice,
before and after the fixative addition.
Statistical analysis and software. Analysis of the leaching data was performed at
several levels of granularity using SPSS Statistics (IBM, version 26). Two and one-
way ANOVAs were used to assess significant differences between gravity condi-
tions, organisms, ground and space samples, and between controls, in combina-
tions described in the results. In these analyses, data across all REEs was used
(the tests did not discriminate REEs). Data were log
10
transformed and tests for
normality of data and equal variances (Levene’s tests) were carried out. Tukey tests
were performed where appropriate to examine pairwise comparisons.
To investigate differences between gravity conditions and organisms or controls
for specific REEs, a two-sample independent two-tailed Student’sttest was used
between pairs of conditions for specific REEs, accepting that the small sample sizes
make these tests less reliable than the aggregate ANOVA analyses.
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w ARTICLE
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 9
Content courtesy of Springer Nature, terms of use apply. Rights reserved
RStudio 1.2.5033 was used to analyse and visualise the ground pH experiment.
Microsoft Excel (2016) was used to collect data and Microsoft Word (2016) was
used to prepare the manuscript and associated text files.
Data availability
The presented data in the paper are available on the Edinburgh Datashare repository at
https://doi.org/10.7488/ds/2908 and from the corresponding author.
Received: 3 July 2020; Accepted: 7 October 2020;
References
1. Druschel, G. K. & Kappler, A. Geomicrobiology and microbial geochemistry.
Elements 11, 389–394 (2015).
2. Taunton, A. E., Welch, S. A. & Banfield, J. E. Microbial controls on phosphate
and lanthanide distributions during granite weathering and soil formation.
Chem. Geol. 169, 371–382 (2000).
3. Schulz, S. et al. The role of microorganisms at different stages of ecosystem
development for soil formation. Biogeosci.10, 3983–3996 (2013).
4. Kalev, S. D. & Toor, G. S. in Green Chemistry: An Inclusive Approach (eds
Torok, B. & Dransfield, T.) 339–357 (Elsevier, 2018).
5. Rawlings, D. E. & Silver, S. Mining with microbes. Nat. Biotechnol. 13,
773–778 (1995).
6. Johnson, D. B. Biomining: biotechnologies for extracting and recovering
metals from ores and waste materials. Curr. Opin. Biotechnol. 30,24–31
(2014).
7. Das, T., Ayyappan, S. & Chaudhury, G. R. Factors affecting bioleaching
kinetics of sulfide ores using acidophilic micro-organisms. Biometals 12,1–10
(1999).
8. Hong, Y. & Valix, M. Bioleaching of electronic waste using acidophilic sulfur
oxidising bacteria. J. Clean. Prod. 65, 564–472 (2014).
9. Hilson, G. & Monhemius, A. J. Alternatives to cyanide in the gold mining
industry: what prospects for the future? J. Clean. Prod. 14, 1158–1167 (2006).
10. Gu, T., Rastegar, S. O., Mousavi, S. M., Li, M. & Zhou, M. Advances in
bioleaching for recovery of metals and bioremediation of fuel ash and sewage
sludge. Biores. Technol. 261, 428–440 (2018).
11. Reed, D. W., Fujita, Y., Daubaras, D. L., Jiao, Y. & Thompson, V. S.
Bioleaching of rare earth elements from waste phosphors and cracking
catalysts. Hydrometallurgy 166,34–40 (2016).
12. Bosecker, K. Bioleaching: metal solubilization by microorganisms. FEMS
Microbiol. Rev. 20, 591–604 (1997).
13. Rezza, I., Salinas, E., Elorza, M., Sanz de Tosetti, M. & Donati, E. Mechanisms
involved in bioleaching of an aluminosilicate by heterotrophic
microorganisms. Process Biochem. 36, 495–500 (2001).
14. Schippers, A. et al. Biomining: metal recovery from ores with microorganisms.
Adv. Biochem. Eng. Biotechnol. 141,1–47 (2014).
15. Sklodowska, A. & Matlakowska, R. in Microbial Processing of Metal Sulfides
(eds Donati, E. R. & Sand, W.) 121–129 (Springer, 2007).
16. Massari, S. & Ruberti, M. Rare earth elements as critical raw materials: focus
on international markets and future strategies. Res. Policy 38,36–43 (2013).
17. Keltjens, J. T., Pol, A., Reimann, J. & Op den Camp, H. J. M. PQQ-dependent
methanol dehydrogenases: rare-earth elements make a difference. Appl.
Microbiol. Biotechnol. 98, 6163–6183 (2014).
18. Huang, J. et al. Rare earth element alcohol dehydrogenases widely occur
among globally distributed, numerically abundant and environmentally
important microbes. ISME J. 13, 2005–2017 (2019).
19. Pol, A. et al. Rare earth metals are essential for methanotrophic life in volcanic
mudpots. Environ. Microbiol. 16, 255–264 (2014).
20. Barmettler, F., Castelberg, C., Fabbri, C. & Brandl, H. Microbial mobilization
of rare earth elements (REE) from mineral solids—a mini review. AIMS
Microbiol. 2, 190–204 (2016).
21. Chen, B. et al. An experimental study on the effects of microbes on the
migration and accumulation of REE in the weathering crust of granite. Chin. J.
Geochem. 19, art. 280 (2000).
22. Cockell, C. S. Geomicrobiology beyond Earth—Microbe-mineral interactions
in space exploration and settlement. Trends Microbiol. 18, 308–314 (2010).
23. Cockell, C. S. Synthetic geomicrobiology: engineering microbe-mineral
interactions for space exploration and settlement. Int. J. Astrobiol. 10, 315–324
(2011).
24. Montague, M. et al. The role of synthetic biology for In Situ Resource
Utilization (ISRU). Astrobiology 12, 1135–1142 (2012).
25. Liu, Y. D. et al. Control of Lunar and Martian dust—experimental insights
from artificial and natural cyanobacterial and algal crusts in the desert of
Inner Mongolia, China. Astrobiology 8,75–86 (2008).
26. Mautner, M. N. Biological potential of extraterrestrial materials—1. Nutrients
in carbonaceous meteorites, and effects on biological growth. Planet. Space Sci.
45, 653–661 (1997).
27. Rothschild, L. J. Synthetic biology meets bioprinting: enabling technologies
for humans on Mars (and Earth). Biochem. Soc. Trans. 44,1158–1164
(2016).
28. Horneck, G., Klaus, D. M. & Mancinelli, R. L. Space microbiology. Microbiol.
Mol. Biol. Rev. 74, 121–156 (2010).
29. Moissl-Eichinger, C., Cockell, C. S. & Rettberg, P. Venturing into new realms?
Microorganisms in space. FEMS Microbiol. Rev. 40, 722–737 (2016).
30. Huang, B., Li, D.-G., Huang, Y. & Liu, C.-T. Effects of spaceflight and
simulated microgravity on microbial growth and secondary metabolism. Mil.
Med. Res. 5, 18 (2018).
31. Santomartino, R. et al. No effect of microgravity and simulated Mars gravity
on final bacterial cell concentrations on the International Space Station:
applications to space bioproduction. Fronti. Micriobiol.11, 579156 (2020).
32. Gasset, G. et al. Growth and division of Escherichia coli under microgravity
conditions. Res. Microbiol. 145, 111–120 (1994).
33. Kacena, M. A. et al. Bacterial growth in space flight: logistic growth curve
parameters for Escherichia coli and Bacillus subtilis.Appl. Microbiol.
Biotechnol. 51, 229–234 (1999).
34. Leys, N. M. E. J., Hendrickx, L., De Boever, P., Baatout, S. & Mergeay, M.
Space flight effects on bacterial physiology. J. Biol. Regul. Homeo. Agents 18,
193–199 (2004).
35. Crabbe, A. et al. Spaceflight enhances cell aggregation and random budding in
Candida albicans.PLoS ONE 8, e80677 (2013).
36. Ruzicka, A., Snyder, G. A. & Taylor, L. A. Comparative geochemistry of
basalts from the Moon, Earth, HED asteroid, and Mars: implications for the
origin of the Moon. Geochim. Cosmochim. Acta 65, 979–997 (2001).
37. McMahon, S., Parnell, J., Ponicka, J., Hole, M. & Boyce, A. The habitability of
vesicles. in martian basalt. Astron. Geophys. 54,1.17–1.21 (2013).
38. McSween, H. Y., Taylor, G. J. & Wyatt, M. B. Elemental composition of the
martian crust. Science 324, 736–739 (2009).
39. McLean, R. J. C., Cassanto, J. M., Barnes, M. B. & Koo, J. H. Bacterial biofilm
formation under microgravity conditions. FEMS Microbiol. Lett. 195, 115–119
(2001).
40. Kim, W. et al. Spaceflight promotes biofilm formation by Pseudomonas
aeruginosa.PLoS ONE 8, e62437 (2013).
41. Pogliani, C. & Donati, E. The role of exopolymers in the bioleaching of a non-
ferrous metal sulphide. J. Indus. Microbiol. Biotechnol. 22,88–92 (1999).
42. Welch, S. A., Barker, W. W. & Banfield, J. F. Microbial extracellular
polysaccharides and plagioclase dissolution. Geochim. Cosmochim. Acta 63,
1405–1419 (1999).
43. Takahashi, Y., Hirata, T., Shimizu, H., Ozaki, T. & Fortin, D. A rare earth
element signature of bacteria in natural waters? Chem. Geol. 244, 569–583
(2007).
44. White, A. F. et al. The effect of temperature on experimental and natural
chemical weathering rates of granitoid rocks. Geochim. Cosmochim. Acta 63,
3277–3291 (1999).
45. Rozas, E. E. et al. Bioleaching of electronic waste using bacteria isolated from
the marine sponge Hymeniacidon heliophila (Porifera). J. Hazard. Mater. 329,
120–130 (2017).
46. Giese, E. C., Carpen, H. L., Bertolino, L. C. & Schneider, C. L. Characterization
and bioleaching of nickel laterite ore using Bacillus subtilis strain. Biotechnol.
Prog. 35, e2860 (2019).
47. Takahashi, Y., Châtellier, X., Hattori, K. H., Kato, K. & Fortin, D. Adsorption
of rare earth elements onto bacterial cell walls and its implication for REE
sorption onto natural microbial mats. Chem. Geol. 219,53–67 (2005).
48. Kucuker, M. A., Xu, X. & Kuchta, K. in Cascade Use in Technologies 2018 (eds
Pehlken, A., Kalverkamp, M. & Wittstock, R.) 45–50 (Springer, 2019).
49. Byloos, B. et al. The impact of space flight on survival and interaction of
Cupriavidus metallidurans CH34 with basalt, a volcanic Moon analog rock.
Front. Microbiol. 8, 671 (2017).
50. Takahashi, Y. et al. A new method for the determination of CeIII/CeIV ratios
in geological materials; application for weathering, sedimentary, and
diagenetic processes. Earth Planet. Sci. Lett. 182, 201–207 (2000).
51. Xhang, L. et al. Bioleaching of rare earth elements from bastnaesite-bearing
rock by actinobacteria. Chem. Geol. 483, 544–557 (2018).
52. Rasoulnia, P., Barthen, R. & Lakaniemi, A.-M. A critical review of bioleaching
of rare earth elements: the mechanisms and effect of process parameters. Crit.
Rev. Env. Sci. Technol.https://doi.org/10.1080/10643389.2020.1727718 (2020).
53. Willscher, S. & Bosecker, K. Studies on the leaching behaviour of
heterotrophic microorganisms isolated from an alkaline slag dump. Hydromet
71, 257–264 (2003).
54. Lapanje, A., Wimmersberger, C., Furrer, G., Brunner, I. & Frey, B. Pattern of
elemental release during the granite dissolution can be changed by aerobic
heterotrophic bacterial strains isolated from Damma Glacier (Central Alps)
deglaciated granite sand. Soil Microbiol. 63, 865–882 (2011).
ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w
10 NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications
Content courtesy of Springer Nature, terms of use apply. Rights reserved
55. McLeod, C. L. & Krekeler, M. P. S. Sources of extraterrestrial rare earth
alements: to the Moon and beyond. Resources 6, 40 (2017).
56. Wieczorek, M. A. & Phillips, R. J. The “Procellarum KREEP Terrane”:
Implications for mare volcanism and lunar evolution. J. Geophys. Res. 105,
20417–20430 (2000).
57. Mautner, M. N., Conner, A. J., Killham, K. & Deamer, D. W. Biological
potential of extraterrestrial materials. Icarus 129, 245–253 (1997).
58. González-Toril, E. et al. Iron meteorites can support the growth of acidophilic
chemolithoautotrophic microorganisms. Astrobiology 5, 406–414 (2005).
59. Gronstal, A. L. et al. Laboratory experiments on the weathering of iron
meteorites and carbonaceous chondrites by iron-oxidising bacteria. Meteorit.
Planet. Sci. 44, 233–248 (2009).
60. Milojevic, T. et al. Exploring the microbial biotransformation of
extraterrestrial material on nanometer scale. Sci. Rep. 9, 18028 (2019).
61. Loudon, C.-M. et al. BioRock: new experiments and hardware to investigate
microbe–mineral interactions in space. Int. J. Astrobiol. 17, 303–313 (2018).
62. Reddy, G. S. N. & Garcia-Pichel, F. Sphingomonas mucosissima sp. nov. and
Sphingomonas desiccabilis sp. nov., from biological soil crusts in the Colorado
Plateau, US. Int. J. Syst. Evol. Microbiol 57, 1028–1034 (2007).
63. Song, W., Ogawab, N., Oguchic, C. T., Hattad, T. & Matsukuraa, Y. Effect of
Bacillus subtilis on granite weathering: a laboratory experiment. CATENA 70,
275–281 (2007).
64. Diels, L. & Mergeay, M. DNA probe-mediated detection of resistant bacteria
from soils highly polluted by heavy metals. Appl. Environ. Microbiol 56,
1485–1491 (1990).
65. Brim, H. et al. Amplified rDNA restriction analysis and further genotypic
characterisation of metal-resistant soil bacteria and related facultative
hydrogenotrophs. Syst. Appl. Microbiol. 22, 258–268 (1999).
66. Goris, J. et al. Classification of metal-resistant bacteria from industrial
biotopes as Ralstonia campinensis sp. nov., Ralstonia metallidurans sp. nov.
and Ralstonia basilensis Steinle et al. 1998 emend. Int. J. Syst. Evol. Microbiol.
51, 1773–1782 (2001).
67. Sahl, J. W. et al. Subsurface microbial diversity in deep-granitic-fracture water
in Colorado. Appl. Environ. Microbiol. 74, 143–152 (2008).
68. Kelly, L. et al. Bacterial diversity of terrestrial crystalline volcanic rocks,
Iceland. Microb. Ecol. 62,69–79 (2011).
69. Mijnendonckx, K. et al. Characterization of the survival ability of Cupriavidus
metallidurans and Ralstonia pickettii from space-related environments.
Microb. Ecol. 65, 347–360 (2013).
70. Leys, N. et al. The response of Cupriavidus metallidurans CH34 to spaceflight in
the International Space Station. Antonie Van. Leeuwenhoek 96,227–245 (2009).
71. Reasoner, D. J. & Geldreich, E. E. A new medium for the enumeration and
subculture of bacteria from potable water. Appl. Environ. Microbiol 49,1–7
(1985).
72. Fuchs, F. M., Driks, A., Setlow, P. & Moeller, R. An improved protocol for
harvesting Bacillus subtilis colony biofilms. J. Microbiol. Methods 134,7–13
(2017).
Acknowledgements
C.S.C., R.S. and the preparation of the experiment and post-flight analysis were funded
by UK Science and Technology Facilities Council under grant ST/R000875/1. AW was
supported by a Principal’s Career Development PhD Scholarship. R.M., F.M.F. and P.R.
were supported by the DLR grant “DLR-FuE-Projekt ISS LIFE, Programm RF-FuW,
Teilprogramm 475”. F.M.F. was also supported by the Helmholtz Space Life Sciences
Research School at DLR. R.V.H. and N.L. received financial support for this study from
Belspo and ESA through the PRODEX EGEM/Biorock project contract (PEA
4000011082). We thank Laetitia Pichevin for ICP-MS analysis of the basalt substrate. We
thank the European Space Agency (ESA) for offering the flight opportunity. A special
thanks to the dedicated ESA/ESTEC teams, Kayser Italia s.r.l., and the USOC BIOTESC
for the development, integration and operation effort. We are thankful to the UK Space
Agency (UKSA) for the national support to the project, NASA Kennedy for their support
in the experiment integration prior to the SpaceX Falcon 9 CSR-18 rocket launch,
particularly Kamber Scott and Anne Currin, and NASA Ames for hosting the ground
control experiment. We thank SpaceX and Elon Musk for launching our mining
experiment into space.
Author contributions
C.S.C. conceived the BioRock experiment in the framework of the ESA topical team
Geomicrobiology for Space Settlement and Exploration (GESSE). C.S.C., R.S. and K.F.
designed the experiments for this paper. N.N. and C.M.L. carried out ground experiments
and studies in preparation for flight. C.S.C., R.S. and A.C.W. integrated the hardware for
spaceflight and ground controls. C.S.C., R.S. and L.J.E. produced the experimental data.
C.S.C. and R.S. performed the data analyses. R.M., P.R., F.F. and R.V.H., N.L., I.C. pro-
vided B. subtilis and C. metallidurans samples, respectively. L.P. performed the procedures
on-board the ISS. R.D., J.H., J.K., A.K., N.C. and L.Z. supervised the technical organisation
and implementation of the experiment at ESA. J.D.W., M.H. and B.R. supervised the flight
procedures. A.M., S.P., F.C., G.L., M.B. and V.Z. designed and fabricated the hardware.
R.C.E. and J.W. hosted the ground control experiment. C.S.C. wrote the manuscript.
All authors discussed the results and commented on the manuscript. C.S.C. and R.S.
contributed equally to the work.
Competing interests
Authors V.Z., M.B., A.M., S.S.P., F.C. and G.L. were employed by the company Kayser
Italia S.r.l. The remaining authors declare no competing interests.
Additional information
Supplementary information is available for this paper at https://doi.org/10.1038/s41467-
020-19276-w.
Correspondence and requests for materials should be addressed to C.S.C.
Peer review information Nature Communications thanks Anna Kaksonen, Robert
McLean, and Elizabeth Watkin for their contributions to the peer review of this work.
Peer review reports are available.
Reprints and permission information is available at http://www.nature.com/reprints
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative
Commons license, and indicate if changes were made. The images or other third party
material in this article are included in the article’s Creative Commons license, unless
indicated otherwise in a credit line to the material. If material is not included in the
article’s Creative Commons license and your intended use is not permitted by statutory
regulation or exceeds the permitted use, you will need to obtain permission directly from
the copyright holder. To view a copy of this license, visit http://creativecommons.org/
licenses/by/4.0/.
© The Author(s) 2020
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-19276-w ARTICLE
NATURE COMMUNICATIONS | (2020) 11:5523 | https://doi.org/10.1038/s41467-020-19276-w | www.nature.com/naturecommunications 11
Content courtesy of Springer Nature, terms of use apply. Rights reserved
1.
2.
3.
4.
5.
6.
Terms and Conditions
Springer Nature journal content, brought to you courtesy of Springer Nature Customer Service Center GmbH (“Springer Nature”).
Springer Nature supports a reasonable amount of sharing of research papers by authors, subscribers and authorised users (“Users”), for small-
scale personal, non-commercial use provided that all copyright, trade and service marks and other proprietary notices are maintained. By
accessing, sharing, receiving or otherwise using the Springer Nature journal content you agree to these terms of use (“Terms”). For these
purposes, Springer Nature considers academic use (by researchers and students) to be non-commercial.
These Terms are supplementary and will apply in addition to any applicable website terms and conditions, a relevant site licence or a personal
subscription. These Terms will prevail over any conflict or ambiguity with regards to the relevant terms, a site licence or a personal subscription
(to the extent of the conflict or ambiguity only). For Creative Commons-licensed articles, the terms of the Creative Commons license used will
apply.
We collect and use personal data to provide access to the Springer Nature journal content. We may also use these personal data internally within
ResearchGate and Springer Nature and as agreed share it, in an anonymised way, for purposes of tracking, analysis and reporting. We will not
otherwise disclose your personal data outside the ResearchGate or the Springer Nature group of companies unless we have your permission as
detailed in the Privacy Policy.
While Users may use the Springer Nature journal content for small scale, personal non-commercial use, it is important to note that Users may
not:
use such content for the purpose of providing other users with access on a regular or large scale basis or as a means to circumvent access
control;
use such content where to do so would be considered a criminal or statutory offence in any jurisdiction, or gives rise to civil liability, or is
otherwise unlawful;
falsely or misleadingly imply or suggest endorsement, approval , sponsorship, or association unless explicitly agreed to by Springer Nature in
writing;
use bots or other automated methods to access the content or redirect messages
override any security feature or exclusionary protocol; or
share the content in order to create substitute for Springer Nature products or services or a systematic database of Springer Nature journal
content.
In line with the restriction against commercial use, Springer Nature does not permit the creation of a product or service that creates revenue,
royalties, rent or income from our content or its inclusion as part of a paid for service or for other commercial gain. Springer Nature journal
content cannot be used for inter-library loans and librarians may not upload Springer Nature journal content on a large scale into their, or any
other, institutional repository.
These terms of use are reviewed regularly and may be amended at any time. Springer Nature is not obligated to publish any information or
content on this website and may remove it or features or functionality at our sole discretion, at any time with or without notice. Springer Nature
may revoke this licence to you at any time and remove access to any copies of the Springer Nature journal content which have been saved.
To the fullest extent permitted by law, Springer Nature makes no warranties, representations or guarantees to Users, either express or implied
with respect to the Springer nature journal content and all parties disclaim and waive any implied warranties or warranties imposed by law,
including merchantability or fitness for any particular purpose.
Please note that these rights do not automatically extend to content, data or other material published by Springer Nature that may be licensed
from third parties.
If you would like to use or distribute our Springer Nature journal content to a wider audience or on a regular basis or in any other manner not
expressly permitted by these Terms, please contact Springer Nature at
onlineservice@springernature.com
Content uploaded by Rosa Santomartino
Author content
All content in this area was uploaded by Rosa Santomartino on Nov 13, 2020
Content may be subject to copyright.
Content uploaded by Charles S Cockell
Author content
All content in this area was uploaded by Charles S Cockell on Nov 10, 2020
Content may be subject to copyright.